Determination of ketamine and xylazine in rat brain by liquid-liquid extraction and gas chromatography-mass spectrometry

Introduction: Ketamine and xylazine are injectable drugs used in human or veterinary medicine mainly in combination for inducing/starting then maintaining general anesthesia; they also provide relief of pain and distress.
Methods: The paper presents a method for determining nanogram quantities of ketamine and xylazine in rat brain using liquid-liquid extractions and gas chromatography-mass spectrometry in selective ion monitoring mode. This technique requires only 0.5 g of sample, and xylazine d6 as an internal standard.
Results: The method was found to be linear between 1 and 40 µg/g of brain. Limits of quantification were 425 and 114 ng (approximately 0.85 and 0.23 µg/g of brain) for ketamine and xylazine respectively. The reliability of the method in terms of accuracy, within-day and between-day precisions has been also demonstrated. For xylazine, bias and intra-day precision were below 3.0 % while between-day precision was below 10.5 %; in the same manner, the values for ketamine were below 7 %, 11.1% and 20.9%, respectively. In addition, the stability of the analytes in the matrix at -80 °C has been checked over five months and stated for at least 1 month in the worst conditions.
Discussion: The present procedure was successfully applied to animals to determine (for the first time) the in vivo brain levels of both drugs following systemic administrations.